Abstract Background. Traumatic injury accounts for an abysmal amount morbidity and mortality, much of which is secondary to hemorrhage. For patients that are injured, major bleeding accounts for the largest number of deaths that are potentially preventable both in military and civilian trauma. Thus, the development of adjuncts to standard care and resuscitation that can be instituted in the field early in the care of these civilians and active duty soldiers has the potential to significantly improve outcomes. Our investigations have focused on the understanding of the role of carbon monoxide (CO) in physiological and pathophysiological conditions, as well as the development of CO as a therapeutic. CO is produced endogenously in the breakdown of heme by heme oxygenase enzymes and has been shown to possess significant anti-inflammatory properties. Exogenous CO can be harnessed for its cytoprotective properties and we have been studying the use of inhaled CO as a therapeutic in pre-clinical models since 1999. In an effort to develop a life-saving resuscitation adjunct/biologic for the treatment of trauma victims we have developed the following hypothesis: Carbon Monoxide protects against the development of shock, circulatory failure and death from hemorrhage/trauma. The goal of this application is to study the use of CO as a therapeutic agent and resuscitative adjunct in the treatment of hemorrhagic shock in pre-clinical models. Our strong preclinical preliminary data demonstrates that inhaled CO can protect against the development of shock, inflammation, organ injury, and death from hemorrhage and preliminary studies have been performed illustrating the feasibility of inhaled CO administration in humans. SPECIFIC OBJECTIVE I. To determine the optimum dosing regimen/preparation of CO to protect against the development of shock and circulatory collapse from hemorrhage and trauma in a mouse model. SPECIFIC OBJECTIVE II. To determine the role and mechanism(s) of heme oxygenase/carbon monoxide in protecting against endothelial injury and the development shock and circulatory collapse from hemorrhage. Study Design: In order to study the influence of CO on hemorrhage-induced shock, and death, we will utilize a well-established murine model of hemorrhage. This model will allow thorough investigation of dosing and kinetics of CO for the treatment of hemorrhage. Inhaled CO (25-500 ppm) or pharmacological CO- releasing molecules will be initiated as a therapy at time points relevant to the care of patients with combat injuries. All appropriate controls including sham animals will be included in all investigations. Endpoints will be examined, including time to the development of circulatory collapse and death, as well as clinical measurements of shock such as pH, base deficit, lactate, and coagulation studies. Furthermore, tissues and serum will be collected for determination of organ injury and inflammation. Additionally, the influence of CO and heme oxygenase enzymes on endothelial injury will be investigated. These studies will utilize the murine in vivo model of hemorrhagic shock as well as an in vitro model of hypoxia and inflammatory stimulation in endothelial cells. Investigations will focus on endothelial activation. Studies investigating the mechanisms of action of CO will be executed, focusing on mitogen activated protein kinases. Together, these studies will further our understanding of hemorrhagic shock and have great potential in the development of a possible therapeutic adjunct to improve outcomes of the lives of veterans, active duty soldiers and all civilians. These hypotheses will be tested by addressing the following aims: PUBLIC HEALTH RELEVANCE: Narrative These studies investigating carbon monoxide as a therapeutic to prevent shock and death from hemorrhage have the potential for an outstanding impact on both our understanding of the consequences of hemorrhage and in the clinical care of trauma patients. Hemorrhage continues to account for a significant proportion of the morbidity and mortality in victims of trauma. Additionally, exsanguination is the leading cause of possibly preventable death in both civilian and military trauma. We currently lack effective therapeutic adjuncts to prevent early deaths from the development of circulatory collapse and late deaths from the development of multiple organ dysfunction and sepsis. Given the magnitude of this clinical problem, as well as the deficit in current therapeutics, this proposal is highly responsive to the health care needs of the Armed Forces, the U.S. Veteran population, and the general public.